The fluid medium which supports the axenic growth of Entamoeba histolyica will be simplified by the omission of non-essential factors and by the substitution of chemically defined substances for the complex natural products which it contains. In determining the required concentrations of essential nutrients the criterion used will be rate of growth (cell counts and measured cell mass) plus the ability to sustain growth for a minimum of 32 generations in serial culture. Information on pinocytotic rate and specific membrane transport systems will be developed and employed to estimate the necessary concentrations of the individual precursors of protein and ribonucleic acid synthesis. The objective is to achieve ameba growth in a chemically defined medium. The major components of currently employed axenic growth medium, serum, enzymic casein digest, enzymic liver digest (or yeast extract) will investigated by fractionation or substitution procedures. An intermediate objective will be to identify those factors which lead to maximum rate of growth and cell harvest in medium containing complex natural products and to supplement the medium with them to avoid the variability inherent in the use of different batches of the natural products.